Providing drug depots for sustained-release action is essential for efficient treatment of patients which require a steady administration of specific pharmaceutically active substances, especially if the drug is desired to be applied inside the body. A prerequisite for an adequate drug depot is that the release of the pharmaceutically active substance from such a depot is controllable by specific retardation processes. This implies that the pharmaceutically active substance has to be connected to the depot matrix either in a reversible or an irreversible way.
The matrix to which the pharmaceutically active substance is bound should not only have an affinity to the active substance but also have biocompatible properties. Preferably, such depot matrices are biodegradable within the body of the patient so that no further treatment of the patient for removing the emptied depot is necessary.
Because of their advantageous biological properties especially fibrin gels have been proposed as preferred drug depot matrices (see e.g. AT 369 990). Fibrin gels are easy to prepare, have good biocompatibility and their biological degradation inside the body can be regulated. However, due to the hydrated and wide porous structure of fibrin, diffusion of pharmaceutically active substances occurs with a rate much too fast for most purposes even if the fibrin gel is highly cross-linked by excess addition of its natural cross-linking effector, factor XIII.
In preliminary experiments carried out for the present invention it could be shown that different proteins, such as β-galactosidase are completely released from a fibrin gel within three days or less.
It has therefore been proposed to covalently link bioactive factors to a fibrin network by linking a transglutaminase substrate domain to a bioactive factor using factor XIIIa activity (WO98/43686). However, covalent binding of the drug of interest to the fibrin network may result in a binding too strong to allow a sufficient release of the drug to the patient. Not all drugs are compatible with covalent binding. Moreover, the fibrin clot might become unstable because cross-linking sites are used by the transglutaminase substrates, which are essential for this reaction.
It is therefore an object to provide a drug depot having satisfactory biocompatibility and a regulatable half-life in the patient's body.
It is a further object of the present invention to provide for an alternative drug depot based on fibrin, especially without altering the active substance or using enzymatic activity for its linkage.
Another object of the present invention is to provide a drug depot with capacity for efficient retardation of diffusion of the biologically active substance compared to the release time of this drug by diffusion from a standard fibrin gel. cl SUMMARY OF THE INVENTION
These objects are solved by a fibrin/fibrinogen-binding conjugate comprising a fibrin/fibrinogen-binding moiety, a substance capturing moiety capable of reversibly binding to a pharmaceutically active substance, and a pharmaceutically active substance, wherein said fibrin/fibrinogen-binding moiety is bound to said substance capturing moiety.
With the conjugate according to the present invention, binding partners with affinity to fibrin or fibrinogen are used to link binding partners of pharmaceutically active substances to a fibrin gel. Due to these fibrin or fibrinogen binding moieties, the conjugates are bound sufficiently to the fibrin matrix so that elution of the pharmaceutically active substance is not possible by simple diffusion, but mainly dependent on the affinity of the fibrin/fibrinogen-binding moiety to fibrin and on the binding affinity of the substance capturing moiety to the pharmaceutically active substance.
The binding of the fibrin/fibrinogen-binding moiety to the substance capturing moiety can be covalently, e.g. using chemical linkers such as 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) known in the art, or by electrostatic forces.
In another embodiment of the invention, the conjugate is a fusion protein comprising a fibrin/fibrinogen binding moiety covalently linked to a pharmaceutically active, substance, without an intervening substance capturing moiety.
The term “fibrin/fibrinogen-binding moiety” relates to a binding moiety which is capable of binding either to (1) fibrin or to (2) both fibrinogen and fibrin. If the binding capacity is for both fibrin and fibrinogen, it is possible to form the fibrin gel with fibrinogen molecules which are already “loaded” with the present conjugates to allow a homogeneous formation of the drug depot and a homogeneous distribution of the conjugate, and therefore the pharmaceutical throughout the fibrin drug depot.